The long term objectives of this research are to examine at the molecular level the regulation of nitrogen utilization by the gram negative soil microorganism, Rhizobium fredii. This group of bacteria is relatively newly discovered (1982) and although some data have been obtained, complete characterization of their biochemistry has not been undertaken. For these studies the histidine utilization (hut) genes of R. fredii, will be used as a model system to understand how it can utilize poor nitrogen sources. The specific aims of this application are to: (1) Determine the number and the activity of the R. fredii hut enzymes. (2) Identify the number and locations of hut specific transcriptional starts. (3) Begin DNA sequence analysis of putative regulatory sites upstream of the transcriptional start sites. (4) Construct GUS (beta-Glucuronidase) fusions with cloned R. fredii hut DNA for use in regulation studies and the isolation of mutant regulatory elements. This project would successfully integrate biochemical and DNA/molecular approaches to characterize this system and provide additional opportunities for detailed investigations into prokaryotic gene regulation.